Abstract
Notch is a transmembrane receptor that controls cell fate decisions during development and tissue homeostasis. Both activation and attenuation of the Notch signal are tightly regulated by endocytosis. The adaptor protein Numb acts as an inhibitor of Notch and is known to function within the intracellular trafficking pathways. However, a role for Numb in regulating Notch trafficking has not been defined. Here we show that mammalian Notch1 is constitutively internalized and trafficked to both recycling and late endosomal compartments, and we demonstrate that changes in Numb expression alter the dynamics of Notch1 trafficking. Overexpression of Numb promotes sorting of Notch1 through late endosomes for degradation, whereas depletion of Numb facilitates Notch1 recycling. Numb mutants that do not interact with the ubiquitin-protein isopeptide ligase, Itch, or that lack motifs important for interaction with endocytic proteins fail to promote Notch1 degradation. Our data suggest that Numb inhibits Notch1 activity by regulating post-endocytic sorting events that lead to Notch1 degradation.
Highlights
The coordinated processing, endocytosis, and trafficking of the Notch receptor and its ligand are important in controlling Notch activation
We showed that Numb binds to the Su(Dx) orthologue Itch and cooperates with Itch to enhance the ubiquitination of membrane-associated Notch1 [32], suggesting that Numb might function in a post-endocytic compartment to regulate Notch trafficking
Numb Endocytic Motifs and Interaction with Itch Are Required to Regulate Notch1 Trafficking—To determine the regions of Numb required for modulating Notch1 trafficking, we examined the effects of the Numb mutant (Nb⌬C) lacking the last 41 amino acids of Numb, which includes the NPF and DPF motifs required for Numb binding to ␣-adaptin, Eps15, and EHD4 [41, 42], and the Nb⌬PTBC mutant, which lacks 88 amino acids within the carboxyl-terminal region of the PTB domain required for binding to the E3 ligase, Itch [32] (Fig. 5A)
Summary
The coordinated processing, endocytosis, and trafficking of the Notch receptor and its ligand are important in controlling Notch activation. In Drosophila both Su(Dx) and dNedd act to limit Notch signaling by regulating post-endocytic sorting of Notch [24, 25]. Together these studies suggest that both entry and trafficking of Notch within the endocytic pathway are important in the regulation of its activity. We showed that Numb binds to the Su(Dx) orthologue Itch and cooperates with Itch to enhance the ubiquitination of membrane-associated Notch1 [32], suggesting that Numb might function in a post-endocytic compartment to regulate Notch trafficking
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
